Activation of Pedunculopontine Tegmental Nucleus Alleviates the Pain Induced by the Lesion of Midbrain Dopaminergic Neurons.

The loss of midbrain dopaminergic (DA) neurons is the fundamental pathological feature of Parkinson's disease (PD). PD causes chronic pain in two-thirds of patients. Recent studies showed that the activation of the pedunculopontine tegmental nucleus (PPTg) can effectively relieve inflammatory pain and neuropathic pain. The PPTg is located in the pontomesencephalic tegmentum, a target of deep brain stimulation (DBS) treatment in PD, and is involved in motor control and sensory integration. To test whether the lesion of midbrain DA neurons induced pain hypersensitivity, and whether the chemogenetic activation of the PPTg could modulate the pain, the AAV-hM3Dq receptor was transfected and expressed into the PPTg neurons of 6-hydroxydopamine-lesioned mice. In this study, von Frey, open field, and adhesive tape removal tests were used to assess animals' pain sensitivity, locomotor activity, and sensorimotor function and somatosensory perception, respectively. Here, we found that the lesion of midbrain DA neurons induced a minor deficit in voluntary movement but did not affect sensorimotor function and somatosensory perception in the tape removal test. The results showed that lesion led to pain hypersensitivity, which could be alleviated both by levodopa and by the chemogenetic activation of the PPTg. Activating the PPTg may be a potential therapeutic strategy to relieve pain phenotypes in PD.

SIRT1 mediates the excitability of spinal CaMKIIα-positive neurons and participates in neuropathic pain by controlling Nav1.3.

AIMS: Neuropathic pain is a common chronic pain disorder, which is largely attributed to spinal central sensitization. Calcium/calmodulin-dependent protein kinase II alpha (CaMKIIα) activation in the spinal dorsal horn (SDH) is a major contributor to spinal sensitization. However, the exact way that CaMKIIα-positive (CaMKIIα+) neurons in the SDH induce neuropathic pain is still unclear. This study aimed to explore the role of spinal CaMKIIα+ neurons in neuropathic pain caused by chronic constriction injury (CCI) and investigate the potential epigenetic mechanisms involved in CaMKIIα+ neuron activation. METHODS: CCI-induced neuropathic pain mice model, Sirt1loxP/loxP mice, and chemogenetic virus were used to investigate whether the activation of spinal CaMKIIα+ neurons is involved in neuropathic pain and its involved mechanism. Transcriptome sequence, western blotting, qRT-PCR, and immunofluorescence analysis were performed to assay the expression of related molecules and activation of neurons. Co-immunoprecipitation was used to observe the binding relationship of protein. Chromatin immunoprecipitation (ChIP)-PCR was applied to analyze the acetylation of histone H3 in the Scn3a promoter region. RESULTS: The expression of sodium channel Nav1.3 was increased and the expression of SIRT1 was decreased in the spinal CaMKIIα+ neurons of CCI mice. CaMKIIα neurons became overactive after CCI, and inhibiting their activation relieved CCI-induced pain. Overexpression of SIRT1 reversed the increase of Nav1.3 and alleviated pain, while knockdown of SIRT1 or overexpression of Nav1.3 promoted CaMKIIα+ neuron activation and induced pain. By knocking down spinal SIRT1, the acetylation of histone H3 in the Scn3a (encoding Nav1.3) promoter region was increased, leading to an increased expression of Nav1.3. CONCLUSION: The findings suggest that an aberrant reduction of spinal SIRT1 after nerve injury epigenetically increases Nav1.3, subsequently activating CaMKIIα+ neurons and causing neuropathic pain.

TET1 Participates in Complete Freund's Adjuvant-induced Trigeminal Inflammatory Pain by Regulating Kv7.2 in a Mouse Model.

Trigeminal inflammatory pain is one of the most severe pain-related disorders in humans; however, the underlying mechanisms remain largely unknown. In this study, we investigated the possible contribution of interaction between ten-eleven translocation methylcytosine dioxygenase 1 (TET1) and the voltage-gated K+ channel Kv7.2 (encoded by Kcnq2) to orofacial inflammatory pain in mice. We found that complete Freund's adjuvant (CFA) injection reduced the expression of Kcnq2/Kv7.2 in the trigeminal ganglion (TG) and induced orofacial inflammatory pain. The involvement of Kv7.2 in CFA-induced orofacial pain was further confirmed by Kv7.2 knockdown or overexpression. Moreover, TET1 knockdown in Tet1flox/flox mice significantly reduced the expression of Kv7.2 and M currents in the TG and led to pain-like behaviors. Conversely, TET1 overexpression by lentivirus rescued the CFA-induced decreases of Kcnq2 and M currents and alleviated mechanical allodynia. Our data suggest that TET1 is implicated in CFA-induced trigeminal inflammatory pain by positively regulating Kv7.2 in TG neurons.

Cav3.2 T-Type calcium channels downregulation attenuates bone cancer pain induced by inhibiting IGF-1/HIF-1α signaling pathway in the rat spinal cord.

Background: Bone cancer pain (BCP) is one of the most ubiquitous and refractory symptoms of cancer patients that needs to be urgently addressed. Substantial studies have revealed the pivotal role of Cav3.2 T-type calcium channels in chronic pain, however, its involvement in BCP and the specific molecular mechanism have not been fully elucidated. Methods: The expression levels of Cav3.2, insulin-like growth factor 1(IGF-1), IGF-1 receptor (IGF-1R) and hypoxia-inducible factor-1α (HIF-1α) were detected by Western blot in tissues and cells. X-ray and Micro CT used to detect bone destruction in rats. Immunofluorescence was used to detect protein expression and spatial location in the spinal dorsal horn. Electrophoretic mobility shift assay used to verify the interaction between HIF-1α and Cav3.2. Results: The results showed that the expression of Cav3.2 channel was upregulated and blockade of this channel alleviated mechanical allodynia and thermal hyperalgesia in BCP rats. Additionally, inhibition of IGF-1/IGF-1R signaling not only reversed the BCP-induced upregulation of Cav3.2 and HIF-1α, but also decreased nociceptive hypersensitivity in BCP rats. Inhibition of IGF-1 increased Cav3.2 expression levels, which were abolished by pretreatment with HIF-1α siRNA in PC12 cells. Furthermore, nuclear HIF-1α bound to the promoter of Cav3.2 to regulate the Cav3.2 transcription level, and knockdown of HIF-1α suppresses the IGF-1-induced upregulation of Cav3.2 and pain behaviors in rats with BCP. Conclusion: These findings suggest that spinal Cav3.2 T-type calcium channels play a central role during the development of bone cancer pain in rats via regulation of the IGF-1/IGF-1R/HIF-1α pathway.

Epigenetic mechanisms of Müller glial reprogramming mediating retinal regeneration.

Retinal degenerative diseases, characterized by retinal neuronal death and severe vision loss, affect millions of people worldwide. One of the most promising treatment methods for retinal degenerative diseases is to reprogram non-neuronal cells into stem or progenitor cells, which then have the potential to re-differentiate to replace the dead neurons, thereby promoting retinal regeneration. Müller glia are the major glial cell type and play an important regulatory role in retinal metabolism and retinal cell regeneration. Müller glia can serve as a source of neurogenic progenitor cells in organisms with the ability to regenerate the nervous system. Current evidence points toward the reprogramming process of Müller glia, involving changes in the expression of pluripotent factors and other key signaling molecules that may be regulated by epigenetic mechanisms. This review summarizes recent knowledge of epigenetic modifications involved in the reprogramming process of Müller glia and the subsequent changes to gene expression and the outcomes. In living organisms, epigenetic mechanisms mainly include DNA methylation, histone modification, and microRNA-mediated miRNA degradation, all of which play a crucial role in the reprogramming process of Müller glia. The information presented in this review will improve the understanding of the mechanisms underlying the Müller glial reprogramming process and provide a research basis for the development of Müller glial reprogramming therapy for retinal degenerative diseases.

MAPK-ERK-CREB signaling pathway upregulates Nav1.6 in oxaliplatin-induced neuropathic pain in the rat.

The voltage-gated sodium channel subtype Nav1.6 is involved in the electrophysiological changes of primary sensory neurons that occur in oxaliplatin-induced neuropathic pain, but its regulatory mechanism remains unclear. In this study, Western blot, RT-qPCR, immunofluorescence staining, chromatin immunoprecipitation were used to prove the mechanism of MAPK-ERK-CREB signaling pathway participating in oxaliplatin-induced neuropathic pain by regulating Nav1.6. The results showed that p-Raf1 and p-ERK, key molecules in MAPK/ERK pathway, and Nav1.6 were significantly increased in DRGs of oxaliplatin-induced neuropathic pain rats. Inhibition of p-Raf1 and p-ERK respectively not only reduced the expression of Nav1.6 protein in DRGs of OXA rats, but also caused a decrease in Nav1.6 mRNA, which led us to further explore the transcription factor CREB regulated by MAPK/ERK pathway. Results showed that CREB was co-distributed with Nav1.6. Inhibition of CREB resulted in decreased mRNA and protein expression of Nav1.6, and alleviated oxaliplatin-induced neuropathic pain. A chromatin immunoprecipitation experiment proved that OXA caused p-CREB to directly bind to the promoter region of Scn8A, which is the encoding gene for Nav1.6, and promote the transcription of Scn8A. In summary, in this study, we found that oxaliplatin can activate the MAPK/ERK pathway, which promotes the expression and activation of CREB and leads to an increase in Scn8A transcription, and then leads to an increase in Nav1.6 protein expression to enhance neuronal excitability and cause pain. This study provides an experimental basis for the molecular mechanism of sodium channel regulation in oxaliplatin-induced neuropathic pain.

The role of epigenetic methylation/demethylation in the regulation of retinal photoreceptors.

Photoreceptors are integral and crucial for the retina, as they convert light into electrical signals. Epigenetics plays a vital role in determining the precise expression of genetic information in space and time during the development and maturation of photoreceptors, cell differentiation, degeneration, death, and various pathological processes. Epigenetic regulation has three main manifestations: histone modification, DNA methylation, and RNA-based mechanisms, where methylation is involved in two regulatory mechanisms-histone methylation and DNA methylation. DNA methylation is the most studied form of epigenetic modification, while histone methylation is a relatively stable regulatory mechanism. Evidence suggests that normal methylation regulation is essential for the growth and development of photoreceptors and the maintenance of their functions, while abnormal methylation can lead to many pathological forms of photoreceptors. However, the role of methylation/demethylation in regulating retinal photoreceptors remains unclear. Therefore, this study aims to review the role of methylation/demethylation in regulating photoreceptors in various physiological and pathological situations and discuss the underlying mechanisms involved. Given the critical role of epigenetic regulation in gene expression and cellular differentiation, investigating the specific molecular mechanisms underlying these processes in photoreceptors may provide valuable insights into the pathogenesis of retinal diseases. Moreover, understanding these mechanisms could lead to the development of novel therapies that target the epigenetic machinery, thereby promoting the maintenance of retinal function throughout an individual's lifespan.

Blocking autofluorescence in brain tissues affected by ischemic stroke, hemorrhagic stroke, or traumatic brain injury.

Autofluorescence is frequently observed in animal tissues, interfering with an experimental analysis and leading to inaccurate results. Sudan black B (SBB) is a staining dye widely used in histological studies to eliminate autofluorescence. In this study, our objective was to characterize brain tissue autofluorescence present in three models of acute brain injury, including collagenase-induced intracerebral hemorrhage (ICH), traumatic brain injury (TBI), and middle cerebral artery occlusion, and to establish a simple method to block autofluorescence effectively. Using fluorescence microscopy, we examined autofluorescence in brain sections affected by ICH and TBI. In addition, we optimized a protocol to block autofluorescence with SBB pretreatment and evaluated the reduction in fluorescence intensity. Compared to untreated, pretreatment with SBB reduced brain tissue autofluorescence in the ICH model by 73.68% (FITC), 76.05% (Tx Red), and 71.88% (DAPI), respectively. In the TBI model, the ratio of pretreatment to untreated decreased by 56.85% (FITC), 44.28% (Tx Red), and 46.36% (DAPI), respectively. Furthermore, we tested the applicability of the protocol using immunofluorescence staining or Cyanine-5.5 labeling in the three models. SBB treatment is highly effective and can be applied to immunofluorescence and fluorescence label imaging techniques. SBB pretreatment effectively reduced background fluorescence but did not significantly reduce the specific fluorescence signal and greatly improved the signal-to-noise ratio of fluorescence imaging. In conclusion, the optimized SBB pretreatment protocol blocks brain section autofluorescence of the three acute brain injury models.

Tryptophan Metabolism in Central Nervous System Diseases: Pathophysiology and Potential Therapeutic Strategies.

The metabolism of L-tryptophan (TRP) regulates homeostasis, immunity, and neuronal function. Altered TRP metabolism has been implicated in the pathophysiology of various diseases of the central nervous system. TRP is metabolized through two main pathways, the kynurenine pathway and the methoxyindole pathway. First, TRP is metabolized to kynurenine, then kynurenic acid, quinolinic acid, anthranilic acid, 3-hydroxykynurenine, and finally 3-hydroxyanthranilic acid along the kynurenine pathway. Second, TRP is metabolized to serotonin and melatonin along the methoxyindole pathway. In this review, we summarize the biological properties of key metabolites and their pathogenic functions in 12 disorders of the central nervous system: schizophrenia, bipolar disorder, major depressive disorder, spinal cord injury, traumatic brain injury, ischemic stroke, intracerebral hemorrhage, multiple sclerosis, Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, and Huntington's disease. Furthermore, we summarize preclinical and clinical studies, mainly since 2015, that investigated the metabolic pathway of TRP, focusing on changes in biomarkers of these neurologic disorders, their pathogenic implications, and potential therapeutic strategies targeting this metabolic pathway. This critical, comprehensive, and up-to-date review helps identify promising directions for future preclinical, clinical, and translational research on neuropsychiatric disorders.

Mitophagy-related gene signature predicts prognosis, immune infiltration and chemotherapy sensitivity in colorectal cancer.

BACKGROUND: Mitophagy plays essential role in the development and progression of colorectal cancer (CRC). However, the effect of mitophagy-related genes in CRC remains largely unknown. AIM: To develop a mitophagy-related gene signature to predict the survival, immune infiltration and chemotherapy response of CRC patients. METHODS: Non-negative matrix factorization was used to cluster CRC patients from Gene Expression Omnibus database (GSE39582, GSE17536, and GSE37892) based on mitophagy-related gene expression. The CIBERSORT method was applied for the evaluation of the relative infiltration levels of immune cell types. The performance signature in predicting chemotherapeutic sensitivity was generated using data from the Genomics of Drug Sensitivity in Cancer database. RESULTS: Three clusters with different clinicopathological features and prognosis were identified. Higher enrichment of activated B cells and CD4+ T cells were observed in cluster III patients with the most favorable prognosis. Next, a risk model based on mitophagy-related genes was developed. Patients in training and validation sets were categorized into low-risk and high-risk subgroups. Low risk patients showed significantly better prognosis, higher enrichment of immune activating cells and greater response to chemotherapy (oxaliplatin, irinotecan, and 5-fluorouracil) compared to high-risk patients. Further experiments identified CXCL3 as novel regulator of cell proliferation and mitophagy. CONCLUSION: We revealed the biological roles of mitophagy-related genes in the immune infiltration, and its ability to predict patients' prognosis and response to chemotherapy in CRC. These interesting findings would provide new insight into the therapeutic management of CRC patients.

[Treadmill exercise alleviates neuropathic pain by regulating mitophagy of the anterior cingulate cortex in rats].

This study aimed to investigate the effect of treadmill exercise on neuropathic pain and to determine whether mitophagy of the anterior cingulate cortex (ACC) contributes to exercise-mediated amelioration of neuropathic pain. Chronic constriction injury of the sciatic nerve (CCI) was used to establish a neuropathic pain model in Sprague-Dawley (SD) rats. Von-Frey filaments were used to assess the mechanical paw withdrawal threshold (PWT), and a thermal radiation meter was used to assess the thermal paw withdrawal latency (PWL) in rats. qPCR was used to evaluate the mRNA levels of Pink1, Parkin, Fundc1, and Bnip3. Western blot was used to evaluate the protein levels of PINK1 and PARKIN. To determine the impact of the mitophagy inducer carbonyl cyanide m-chlorophenylhydrazone (CCCP) on pain behaviors in CCI rats, 24 SD rats were randomly divided into CCI drug control group (CCI+Veh group), CCI+CCCP low-dose group (CCI+CCCP0.25), CCI+CCCP medium-dose group (CCI+CCCP2.5), and CCI+CCCP high-dose group (CCI+CCCP5). Pain behaviors were assessed on 0, 1, 3, 5, and 7 days after modeling. To explore whether exercise regulates pain through mitophagy, 24 SD rats were divided into sham, CCI, and CCI+Exercise (CCI+Exe) groups. The rats in the CCI+Exe group underwent 4-week low-moderate treadmill training one week after modeling. The mechanical pain and thermal pain behaviors of the rats in each group were assessed on 0, 7, 14, 21, and 35 days after modeling. Western blot was used to detect the levels of the mitophagy-related proteins PINK1, PARKIN, LC3 II/LC3 I, and P62 in ACC tissues. Transmission electron microscopy was used to observe the ultrastructure of mitochondrial morphology in the ACC. The results showed that: (1) Compared with the sham group, the pain thresholds of the ipsilateral side of the CCI group decreased significantly (P < 0.001). Meanwhile, the mRNA and protein levels of Pink1 were significantly higher, and those of Parkin were lower in the CCI group (P < 0.05). (2) Compared with the CCI+Veh group, each CCCP-dose group showed higher mechanical and thermal pain thresholds, and the levels of PINK1 and LC3 II/LC3 I were elevated significantly (P < 0.05, P < 0.01). (3) The pain thresholds of the CCI+Exe group increased significantly compared with those of the CCI group after treadmill intervention (P < 0.001, P < 0.01). Compared with the CCI group, the protein levels of PINK1 and P62 were decreased (P < 0.001, P < 0.01), and the protein levels of PARKIN and LC3 II/LC3 I were increased in the CCI+Exe group (P < 0.01, P < 0.05). Rod-shaped mitochondria were observed in the ACC of CCI+Exe group, and there were little mitochondrial fragmentation, swelling, or vacuoles. The results suggest that the mitochondrial PINK1/PARKIN autophagy pathway is blocked in the ACC of neuropathic pain model rats. Treadmill exercise could restore mitochondrial homeostasis and relieve neuropathic pain via the PINK1/PARKIN pathway.

Nuclear ATR lysine-tyrosylation protects against heart failure by activating DNA damage response.

Dysregulated amino acid increases the risk for heart failure (HF) via unclear mechanisms. Here, we find that increased plasma tyrosine and phenylalanine levels are associated with HF. Increasing tyrosine or phenylalanine by high-tyrosine or high-phenylalanine chow feeding exacerbates HF phenotypes in transverse aortic constriction and isoproterenol infusion mice models. Knocking down phenylalanine dehydrogenase abolishes the effect of phenylalanine, indicating that phenylalanine functions by converting to tyrosine. Mechanistically, tyrosyl-tRNA synthetase (YARS) binds to ataxia telangiectasia and Rad3-related gene (ATR), catalyzes lysine tyrosylation (K-Tyr) of ATR, and activates the DNA damage response (DDR) in the nucleus. Increased tyrosine inhibits the nuclear localization of YARS, inhibits the ATR-mediated DDR, accumulates DNA damage, and elevates cardiomyocyte apoptosis. Enhancing ATR K-Tyr by overexpressing YARS, restricting tyrosine, or supplementing tyrosinol, a structural analog of tyrosine, promotes YARS nuclear localization and alleviates HF in mice. Our findings implicate facilitating YARS nuclear translocation as a potential preventive and/or interfering measure against HF.

Short-term outcomes of reduced-port laparoscopic surgery versus conventional laparoscopic surgery for total gastrectomy: a single-institute experience.

BACKGROUND: The efficacy of reduced-port laparoscopic surgery (RLS) for total gastrectomy remains unclear. This study focused on evaluating the short-term outcomes of RLS compared with conventional laparoscopic surgery (CLS) for total gastrectomy. METHODS: One hundred and ten patients who underwent completed laparoscopic total gastrectomy for gastric cancer between September 2018 and June 2022 were retrospectively collected and classified into two groups (65 CLS and 45 RLS) according to different operation approach. Twenty-four RLS cases underwent single-incision plus two ports laparoscopic surgery (SILS + 2) and twenty-one underwent single-incision plus one port laparoscopic surgery (SILS + 1). Surgical outcomes, pain intensity, cosmetic and postoperative morbidity, and mortality were compared between groups. RESULTS: The overall incidence of postoperative complications was similar between the CLS group and the RLS group (16.9% vs. 8.9%, P = 0.270). It was also comparable in the Clavien-Dindo classification (P = 0.774). However, compared with the CLS group, the RLS group had a significantly shorter total length of incision (5.6 ± 1.0 cm vs. 7.1 ± 0.7 cm, P = 0.000); shorter time to first ambulation (24.9 ± 5.9 h vs. 27.6 ± 5.0 h, P = 0.009), flatus (3.0 ± 0.8 d vs. 3.5 ± 1.0 d, P = 0.022) and oral intake (4.0 ± 1.6 d vs. 6.1 ± 5.1 d, P = 0.011); lower white blood cell count on the third day after the operation (9.8 ± 4.0*109/L vs. 11.6 ± 4.7*109/L, P = 0.037); and lower visual analogue scale score on postoperative days 1 and 3(3.0 ± 0.7 vs. 3.3 ± 0.7, P = 0.044 and 0.6 ± 0.7 vs. 1.6 ± 0.6, P = 0.000 respectively). On the other hand, it didn't find any difference in short-term outcomes between the SILS + 2 group and the SILS + 1 group (P > 0.05). But the proximal resection margin was longer in the SILS + 2 group than in the SILS + 1 group (2.6 ± 0.7 cm vs. 1.5 ± 0.9 cm, P = 0.046) in patients with adenocarcinoma of the esophagogastric junction (AEG). CONCLUSIONS: RLS for total gastrectomy is a feasible and safe technique when performed by an experienced laparoscopic surgeon. Moreover, compared with SILS + 1, SILS + 2 might have some advantages in AEG patients.

Projections from the Rostral Zona Incerta to the Thalamic Paraventricular Nucleus Mediate Nociceptive Neurotransmission in Mice.

Zona incerta (ZI) is an integrative subthalamic region in nociceptive neurotransmission. Previous studies demonstrated that the rostral ZI (ZIR) is an important gamma-aminobutyric acid-ergic (GABAergic) source to the thalamic paraventricular nucleus (PVT), but whether the ZIR-PVT pathway participates in nociceptive modulation is still unclear. Therefore, our investigation utilized anatomical tracing, fiber photometry, chemogenetic, optogenetic and local pharmacological approaches to investigate the roles of the ZIRGABA+-PVT pathway in nociceptive neurotransmission in mice. We found that projections from the GABAergic neurons in ZIR to PVT were involved in nociceptive neurotransmission. Furthermore, chemogenetic and optogenetic activation of the ZIRGABA+-PVT pathway alleviates pain, whereas inhibiting the activities of the ZIRGABA+-PVT circuit induces mechanical hypersensitivity and partial heat hyperalgesia. Importantly, in vivo pharmacology combined with optogenetics revealed that the GABA-A receptor (GABAAR) is crucial for GABAergic inhibition from ZIR to PVT. Our data suggest that the ZIRGABA+-PVT pathway acts through GABAAR-expressing glutamatergic neurons in PVT mediates nociceptive neurotransmission.

The impact of enhancing publicity and commemoration of body donors at Zhengzhou University, China.

As a fundamental subject of medical education, human anatomy plays a critical role in the development of medical science. However, because of multiple factors including cultural conservativism and limited social understanding, China is facing a particularly severe shortage of bodies donated for anatomy education. Zhengzhou University (ZZU) has continued to uphold whole-body dissection as the preferred method for medical students to learn anatomy. For this study, records of registered individuals (who have signed a body donation agreement) and donors (whose bodies have been received) from 2001 to 2020 were collected and analyzed. The aim of this study was to explore the factors influencing the body donation program (BDP) at ZZU, and then reinforce the social understanding for the BDP. The results showed a significant increase in the numbers of both registered individuals and donors since 2015, which is the year the publicity and commemoration in honor of donors were increased. There were no significant differences between the biological male and female sexes in the registered individuals, but the number of male donors (12.85 ± 10.86, per year) was significantly higher than that of female donors (4.75 ± 4.53, per year). The current donor profile at ZZU is male in his 60/70s, while the profile of registered individuals is male or female in their 60s. Strengthening the publicity and commemoration in honor of donors may contribute to the implementation of BDPs.

Gestational palmitic acid suppresses embryonic GATA-binding protein 4 signaling and causes congenital heart disease.

Dysregulated maternal fatty acid metabolism increases the risk of congenital heart disease (CHD) in offspring with an unknown mechanism, and the effect of folic acid fortification in preventing CHD is controversial. Using gas chromatography coupled to either a flame ionization detector or mass spectrometer (GC-FID/MS) analysis, we find that the palmitic acid (PA) concentration increases significantly in serum samples of pregnant women bearing children with CHD. Feeding pregnant mice with PA increased CHD risk in offspring and cannot be rescued by folic acid supplementation. We further find that PA promotes methionyl-tRNA synthetase (MARS) expression and protein lysine homocysteinylation (K-Hcy) of GATA4 and results in GATA4 inhibition and abnormal heart development. Targeting K-Hcy modification by either genetic ablation of Mars or using N-acetyl-L-cysteine (NAC) decreases CHD onset in high-PA-diet-fed mice. In summary, our work links maternal malnutrition and MARS/K-Hcy with the onset of CHD and provides a potential strategy in preventing CHD by targeting K-Hcy other than folic acid supplementation.

An enriched environment improves long-term functional outcomes in mice after intracerebral hemorrhage by mechanisms that involve the Nrf2/BDNF/glutaminase pathway.

Post-stroke depression exacerbates neurologic deficits and quality of life. Depression after ischemic stroke is known to some extent. However, depression after intracerebral hemorrhage (ICH) is relatively unknown. Increasing evidence shows that exposure to an enriched environment (EE) after cerebral ischemia/reperfusion injury has neuroprotective effects in animal models, but its impact after ICH is unknown. In this study, we investigated the effect of EE on long-term functional outcomes in mice subjected to collagenase-induced striatal ICH. Mice were subjected to ICH with the standard environment (SE) or ICH with EE for 6 h/day (8:00 am-2:00 pm). Depressive, anxiety-like behaviors and cognitive tests were evaluated on day 28 with the sucrose preference test, tail suspension test, forced swim test, light-dark transition experiment, morris water maze, and novel object recognition test. Exposure to EE improved neurologic function, attenuated depressive and anxiety-like behaviors, and promoted spatial learning and memory. These changes were associated with increased expression of transcription factor Nrf2 and brain-derived neurotrophic factor (BDNF) and inhibited glutaminase activity in the perihematomal tissue. However, EE did not change the above behavioral outcomes in Nrf2-/- mice on day 28. Furthermore, exposure to EE did not increase BDNF expression compared to exposure to SE in Nrf2-/- mice on day 28 after ICH. These findings indicate that EE improves long-term outcomes in sensorimotor, emotional, and cognitive behavior after ICH and that the underlying mechanism involves the Nrf2/BDNF/glutaminase pathway.

Spinal Sirtuin 3 Contributes to Electroacupuncture Analgesia in Mice With Chronic Constriction Injury-Induced Neuropathic Pain.

BACKGROUND: Electroacupuncture (EA) is a traditional Chinese therapeutic technique that has a beneficial effect on neuropathic pain; however, the specific mechanism remains unclear. In this study, we investigated whether EA inhibits spinal Ca/calmodulin-dependent protein kinase II (CaMKIIα) phosphorylation through Sirtuin 3 (SIRT3) protein, thus relieving neuropathic pain. MATERIALS AND METHODS: We used wild-type and SIRT3 knockout (SIRT3-/-) mice and used chronic constriction injury (CCI) as a pain model. We performed Western blotting, immunostaining, von Frey, and Hargreaves tests to gather biochemical and behavioral data. Downregulation and overexpression and spinal SIRT3 protein were achieved by intraspinal injection of SIRT3 small interfering RNA and intraspinal injection of lentivirus-SIRT3. To test the hypothesis that CaMKIIα signaling was involved in the analgesic effects of EA, we expressed CaMKIIα-specific designer receptors exclusively activated by designer drugs (DREADDs) in the spinal dorsal horn (SDH) of mice. RESULTS: These results showed that the mechanical and thermal hyperalgesia induced by CCI was related to the decreased spinal SIRT3 expression in the SDH of mice. A significant reduction of mechanical and thermal thresholds was found in the SIRT3-/- mice. SIRT3 overexpression or EA treatment alleviated CCI-induced neuropathic pain and prevented the spinal CaMKIIα phosphorylation. Most importantly, EA increased the expression of spinal SIRT3 protein in the SDH. Downregulation of spinal SIRT3 or CaMKIIα Gq-DREADD activation inhibited the regulatory effect of EA on neuropathic pain. CONCLUSION: Our results showed that CaMKIIα phosphorylation was inhibited by spinal SIRT3 in neuropathic pain and that EA attenuated CCI-induced neuropathic pain mainly by upregulating spinal SIRT3 expression in the SDH of mice.

Dopamine receptor D2, but not D1, mediates the reward circuit from the ventral tegmental area to the central amygdala, which is involved in pain relief.

Pain involves both sensory and affective dimensions. The amygdala is a key player in linking nociceptive stimuli to negative emotional behaviors or affective states. Relief of pain is rewarding and activates brain reward circuits. Whether the reward circuit from the ventral tegmental area (VTA) to the central amygdala (CeA) is involved in pain relief remains unexplored. Using a model of experimental postsurgical pain, we found that pain relief elicited conditioned place preference (CPP), activated CeA-projecting dopaminergic cells in the VTA, and decreased dopaminergic D2 receptor expression in the CeA. Activation of the VTA-CeA neural pathway using optogenetic approaches relieved incisional pain. Administration of a D2 receptor agonist reversed the pain relief elicited by light-induced activation of the VTA-CeA pathway. These findings indicate that the VTA-CeA circuit is involved in pain relief in mice via dopamine receptor D2 in the CeA.